Systematic genetic analysis of Atku70, Atku80 and Atku70xAtku80 mutants in Arabidopsis thaliana

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Althaqafi, Mohammed Mahfouz (2021). Systematic genetic analysis of Atku70, Atku80 and Atku70xAtku80 mutants in Arabidopsis thaliana. University of Birmingham. Ph.D.

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Abstract

Double strands breaks (DSBs) are very serious DNA lesions that can result in large-scale deletions and chromosomal translocations producing genome instability, senescence, cancer or even cell death. In complex eukaryotes, Ku70 and Ku80 interact forming the Ku’s heterodimer which is essential for a DSB repair pathway called canonical Non-Homologous End Joining (cNHEJ). cNHEJ repair pathway is been reported as the main choice for DSBs repair pathway in plants. The Ku heterodimer recognizes and associates directly to the DSBs. This Ku heterodimer will attract the DNA-dependent protein kinases (DNA-PKcs) which will trigger the cNHEJ signalling and activation. Moreover, several studies have also shown that KU70 and KU80 are responsible for further cellular actions such as telomere maintenance and chromosome stability in several organisms including Arabidopsis.

In this study, Atku70 and Atku80 single null homozygous mutants during different generations were analysed systematically. Also, for the first time in Arabidopsis a double Atku70xAtku80 mutant was generated. This genetic analysis has been carried out with plants of the different mutants and genotypes, as well as WT controls growing at the same time and in identical conditions.

Our data suggests that the effect of these mutants increases after several generations of these plants. Some of these effects are decrease plant fertility, some defects in seed germination and hypersensitivity to DNA damages produce by cisplatin. Also, our systematic analysis of AtKu70 and AtKu80 transcripts expression in different Arabidopsis tissues shows different levels of expression and specificity of transcripts among tissues. Interestingly, it seems that the presence of AtKu80 protein have a positive impact in the upregulation of AtKu70 expression levels. Whereas, the absence of Atku70 not only the levels of the different transcripts for AtKu80 genewere affected but also which transcript was expressed on the different tissues. Preliminary cytological analysis in the single and double mutants has shown the presence of anaphase bridges, chromosome miss-segregation and lagging chromosomes during mitotic and meiotic stages as well as telomers lengthening. Most interestingly, we have seen differences between Atku70 and Atku80 and those with the double mutant. Remarkably, although the mean chiasma frequencies observed in the single and double mutants at Metaphase I were not significantly different among them and the WT, univalents and changes in chiasma distribution was observed.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

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